T lymphocytes recognizes antigens through the T cell antigen receptor (TCR) complex. The TCR is a clone-specific heterodimer on T cells, which recognizes its target antigen in association with a major histocompatibility antigen. Moreover, the TCR is highly polymorphic in different T cells. Approximately 90 percent of peripheral blood T cells express a TCR consisting of an .alpha. polypeptide and a .beta. polypeptide and a small percentage of T cells express a TCR consisting of a .gamma. polypeptide and a .delta. polypeptide. See Davis and Bjorkman, 1988, Nature 334:395-402; Marrack and Kappler, 1986, Sci. Amer. 254:36; Meuer et al., 1984, Ann. Rev. Immunol. 2:23-50; Brenner et al., 1986, Nature 322:145-159; Krangel et al., 1987, Science 237:1051-1055; Hata et al., 1987, Science 238:678-682; Hochstenbach et al., 1988, J. Exp. Med. 168:761-776).
The chains of the T cell antigen receptor of a T cell clone are each composed of a unique combination of domains designated variable (V), diversity (D), joining (J), and constant (C) (Siu et al., 1984, Cell 37:393; Yanagi et al., 1985 Proc. Natl. Acad. Sci. USA 82:3430). Hypervariable regions also have been identified (Patten et al., 1984, Nature 312:40; Becker et al., 1985, Nature 317:430). In each T cell clone, the combination of V, D and J domains of both the alpha and the beta chains or both the delta and gamma chains and defines a unique antigen binding site in each T-cell clone. In contrast, the C domain does not participate in antigen binding.
TCR genes, like immunoglobulin genes, consist of regions which arrange during T cell ontogeny (Chien et al., 1984, Nature 312:31-35; Hedrick et al., 1984, Nature 308:149-153; Yanagi et al., 1984, Nature 308:145-149). In genomic DNA, each TCR gene has V, J, and C regions; TCR .beta. and .delta. polypeptides also have D regions. The V, D, J and C regions are separated from one another by spacer regions in the DNA. There are usually many variable region segments and somewhat fewer diversity, junctional, and constant regions segments. As a lymphocyte matures, these various segments are spliced together to create a continuous gene sequence consisting of one V, (D), J, and C regions. TCR diversity, and thus T cell specificity, derives from several sources, (Barth et al, 1985, Nature 316:517-523; Fink et al., 1986, Nature 321:219-225) including: a multiplicity of germline gene segments (Chien et al., 1984, Nature 309:322-326; Malissen et al., 1984, Cell 37:1101-1110; Gascoigne et al., 1984, Nature 310:387-391; Kavaler et al., 1984, Nature 310:421-423; Siu et al., 1984, Nature 311:344-349; Patten et al., 1984, Nature 312:40-46), combinatorial diversity through the assembly of different V, D, J, and C segments (Siu et al., 1984, Cell 37:393-401; Goverman et al., 1985, Cell 40:859-867), and junctional flexibility, N-region diversity and the use of either multiple D regions or any of the three translational reading frames for D.beta. segments. As a result of these mechanisms, TCRs are generated which differ at their N-terminal (called variable, or V regions, constructed from combinations of V, D, and J gene segments) but are the same elsewhere, including their C-terminal (called constant regions). Therefore, an infinite number of TCRs can be established.
The V.beta. gene of the TCR appears to resemble most closely the immunoglobulin V gene in that it has three gene segments, V.beta., D.beta., and J.beta., which rearrange to form a contiguous V.beta. gene (Siu et al., 1984, Cell 37:393-401). The .beta. locus has been well characterized in mice, where it spans 700-800 kilobases of DNA and is comprised of two nearly identical C regions tandemly arranged with one D element and a cluster of 5-6 J elements 5' to each (Kronenberg et al., 1986, Ann Rev. Immunol. 3:537-560). Approximately twenty to thirty V.beta. regions are located upstream (5') to the D, J, and C elements (Behlke et al., 1985, Science, 229:566-570) although V.beta. genes may also be located 3' to the murine C.beta. genes (Malissen et al., 1986, Nature 319:28). Study of the structure and diversity of the human TCR .beta.-chain variable region genes has led to the grouping of genes into district V.beta. subfamilies (Tillinghast et al., 1986, Science 233:879-883; Concannon et al., 1986, Proc. Natl., Acad. Sci. USA 83:6598-6602; Borst et al., 1987, J. Immunol. 139:1952-1959).
The .gamma.TCR gene was identified, first in mice (Saito et al., 1984, Nature 309:757-762; Kranz et al., 1985, Nature 313:762-755; Hayday et al., 1985, Cell 40:259-269) and then in humans (Lefranc et al., 1985, Nature 316:464-466; Murre et al., 1985, Nature 316:549-552). The human .gamma.TCR locus appears to consist of between five and ten variable, five joining, and two constant region genes (Dialynas et al., 1986, Proc. Natl. Acad. Sci. USA 83:2619).
The TCR .alpha. and .delta. locus are adjacent to one another on human chromosome 14. Many TCR .delta. coding segments are located entirely within the .alpha. gene locus (Satyanarayana et al., 1988, Proc. Natl. Acad. Sci. USA 85:8166-8170 Chien et al., 1987, Nature 330:722-727; Elliot et al., 1988, Nature 331:627-631). It is estimated that there are a minimum of 45-50 V.alpha. regions (Becker et al., Nature 317:430-434) whereas there are only approximately 10 V.delta. regions (Chien et al., 1987, supra). Nucleic acid sequences of TCR .alpha. genes have been reported (Sim et al., 1984, Nature 312:771-775; Yanagi et al., 1985, Proc. Natl. Acad. Sci. USA 82:3430-3434; Berkout et al., 1988, Nucl. Acids Res. 16:5208).
Rheumatoid arthritis (RA) is a chronic, recurrent, inflammatory disease primarily involving joints, affecting 1-3% of North Americans. Three times the number of women are afflicted with RA than men. Severe RA patients tend to exhibit extra-articular manifestations including vasculitis, muscle atrophy, subcutaneous nodules, lymphadenopathy, splenomegaly and leukopenia. Spontaneous remission may occur; other patients have brief episodes of acute arthritis with longer periods of low-grade activity; still others progress to severe deformity of joints. It is estimated that about 15% of RA patients become completely incapcitated ("Primer on the Rheumatic Diseases," 8th edition, 1983, Rodman, G. P. & Schumacher, H. R. Eds., Zvaifler, N. J., Assoc. Ed., Arthritis Foundations, Atlanta, Ga.).
The antigenic stimulus initiating the immune response and consequent inflammation is unknown. Certain HLA types (DR4, Dw4, Dw14 and DR1) have an increased prevalence of RA, perhaps leading to a genetic susceptibility to an unidentified factor which initiates the disease process. Relationships between Epstein Barr virus and RA have been suggested.
Many cell types, notably macrophages, synoviocytes and polymorphonuclear leukocytes, participate in the complex inflammatory response which effects joint destruction in R.A. However, a central role for T lymphocytes is suggested by: 1) the rich infiltration of activated T cells at the primary site of RA disease, the synovial tissue van Boxel, J. A., et al., 1975; N. Engl. J. Med., 293:517; Panayi, J. S. et al., 1992, Arthritis Rheum. 35:729; 2) genetic studies linking RA disease susceptibility to a defined amino acid sequence in the third hypervariable region of the DR.beta. chain of the major histocompatibilty complex (MHC) class II molecule P. Gregersen, J. Silver, R. J. Winchester, 1987, Arthritis Rheum. 30:1205); 3) animal models of chronic arthritis in which antigen-specific T cells are capable of transferring disease to naive recipients R. Holmdahl, L. Klareskog, K. Rubin, E. Larsson, H. Wigzell, Scand. J. Immunol. 22:295 (1985); W. van Eden et al., Proc. Natl. Acad. Sci. USA, 1985, 82:5117 (1985)); and 4) amelioration of arthritis, both in murine models of autoimmune disease and in patients with RA, by administration of monoclonal antibody (mAb) reactive with the CD4.sup.+ T cell subset G. E. Rangers, S. Sriram, S. M. Cooper, J. Exp. Med., 1985, 162:11104; G. Horneff, G. R. Burmester, F. Emmrich, J. R. Kalden, Arthritis Rheum., 1991, 34:129).
Previous studies designed to correlate TCR structure with antigen-MHC molecular complex recognition have emphasized the importance of critical amino acid residues in each of the three polymorphic CDR regions of both .alpha. and .beta. chains, with CDR3 playing a dominant role. In both the murine and human systems, T cells specific for a particular peptide--MHC complex often utilize a characteristic amino acid or sequence cluster in the CDR3 region (S. M. Hedrick et al., Science, 1988, 239:1541). Recent studies demonstrate that the introduction of charge altering amino acids in a well defined antigenic peptide results in a T cell response characterized by antigen-specific TCRs which have incorporated reciprocal charge changes in the CDR3 amino acid residues of both .alpha. and .beta. chains (J. L. Jorgensen et al., Nature, 1992, 355:224). This result suggests that these TCR residues bind directly to the antigenic peptide. In a related study, it was found that the murine TCR repertoire recognizing foreign peptides which are highly homologous to self is markedly constrained with respect to TCR V.alpha. and V.beta. gene usage, CDR3 length, and the presence of canonical amino acid residues in the CDR3 domain (J. -L. Casanova et al., J. Exp. Med., 1991, 174:1371). These data suggest that pathogenic T cells mediating autoimmune disease will express TCR which share crucial structural characteristics.
Further support for this hypothesis is found in studies of other autoimmune diseases. Myelin basic protein (MBP) specific T.sub.h cells induce experimental allergic encephalomyelitis (EAE) S. S. Zamvil et al., J. Exp. Med., 1988, 167:1586; J. L. Urban at al., Cell, 1988, 54:577; F. R. Burns et al., J. Exp. Med., 1989, 169:27). Encephalitogenic T cell clones are strongly biased with respect to V.beta. and V.alpha. gene usage as well as CDR3 region structure (D. P Gold et al., J. Immunol., 1992, 148:1712). Recently, it has been shown that TCR V.beta. transcripts isolated from central nervous system lesions of patients with multiple sclerosis (MS) exhibit sequence motifs in the CDR3 region homologous with those expressed by encephalitogenic MBP reactive murine T cell clones (R. Martin et al., J. Exp. Med., 1991, 173:19; J. R. Oksenberg et al., Nature 362, 1993, 68).
While the importance of T cells in RA appears clear, neither the antigen specificity nor the structure of the TCR expressed by disease-inducing T cells has been determined. In an attempt to identify pathogenic T cells among the vast number present in the inflamed joint, investigators have applied molecular techniques to detect T cells which: 1) share TCR structural features, i.e. restricted usage of particular TCR variable gene elements, or 2) are "oligoclonall" with respect to the highly polymorphic antigen binding CDR3 region of the TCR, suggesting antigen-driven expansion at the site of pathology. To date, this approach has yielded conflicting results. Several laboratories have reported evidence of oligoclonality and over-usage of particular TCR V gene products among RA joint-derived T cells (M. D. Howell et al., Proc. Natl. Acad. Sci. USA, 1991, 88:10921; X. Paliard et al., Science, 1991, 253:325; W. V. Williams et al., J. Clin. Invest., 1992, 90:326). However, the TCR V gene families implicated vary from study to study and still other investigations find no evidence for TCR skewing in RA Y. Uematsu et al., Proc. Natl. Acad. Sci. USA, 1991, 88:8534; J. M. van Laar et al., Clin. Exp. Immunol., 1991. 83:353).